Project Summary Ascending infection is hypothesized to underlie a large but unknown fraction of preterm births. The vaginal microbiome represents a physiological barrier to ascending infection through the production of lactic acid by select organisms principally of the Lactobacillus genus. We hypothesize that vaginal microbiota lacking sufficient Lactobacillus spp. (or others), have a heightened susceptibility to ascending infection following the introduction of pathogenic organisms, or the escalation in the relative proportion of seed pathogenic organisms (minor constituents of the normal flora that multiply in permissive states). These susceptible phenotypes may be over-represented among Blacks, or among women practicing specific behaviors that disrupt the normal flora, like douching. We propose to discover what factors influence these unfavorable vaginal microbiome profiles in an existing, diverse cohort of women enrolled in the Pregnancy, Infection and Nutrition (PIN) study. PIN is a landmark longitudinal investigation of the causes of prematurity among Blacks and Whites. Drawing from an existing biobank including vaginal swabs collected at 24-29 weeks' gestation, we propose to: (1) Examine the associations of the vaginal microbiome with spontaneous preterm birth (sPTB), and explore whether these associations differ by maternal race; (2) Examine the influence of innate sPTB risk factors, such as maternal polymorphisms in innate immunity genes and maternal psychosocial stress and depression, on vaginal microbiome profiles overall and stratified by race; and, (3) Examine the influence of exogenous risk factors for sPTB on vaginal microbiome profiles, such as maternal nutritional patterns, maternal smoking, and other health behavior. Furthermore, as an exploratory aim, we will conduct whole genome metagenome sequencing in a subset of PTB cases enriched for an infectious etiology, with delivery before 32 completed weeks' gestation. A better understanding of the relationship between vaginal microbial ecology and premature delivery is poised to offer new opportunities for primary prevention, and the time is ripe to apply these methods to a racially diverse prenatal cohort.